Methanesulfonate Research Articles

HALOGEN FREE [HNMP]+CH3SO3 - IONIC LIQUID MEDIATED BIFUNCTIONALIZED NAPHTHOQUINONE-DIHYDROPYRIMIDIN-(1H)-ONE HYBRIDS AND EVALUATION OF ITS ANTIMICROBIAL POTENTIAL

An efficient halogen free ionic liquid (HFIL) N-methyl-2-pyrrolidonium methane sulfonate [HNMP]+[CH3SO3]- catalyzed Biginelli reaction is reported to synthesize naphthoquinone fused dihydropyrimidin-(1H)-ones (DHPM’s). Diversifying this naphthoquinone constituent provides an access to new and interesting DHPM’s for pharmacological profiling. A series of bifunctionalized naphthoquinone-dihydropyrimidin-(1H)-one conjugates (4a-4l) were synthesized and evaluated for their antimicrobial potential against selected bacterial and fungal pathogenic strains. The resulting naphthoquinone fused dihydropyrimidin-(1H)-one hybrids showed significant activities against all the tested Gram positive (Staphylococcus aureus, Bacillus subtilis) and Gram negative (Escherichia coli, Pseudomonas aeruginosa) bacterial pathogens and fungal strains of Aspergillus niger and Aspergillus flavus. Among the series, the DHPM conjugate 4k>4j>4e>4h samples exhibited highest activities against all the tested strains that proved to be important candidates in the drug development against various life threatening diseases. Presently, one-pot, three-component methodology have been developed for the synthesis of bifunctionalized naphthoquinone linked dihydropyrimidin-(1H)-one are being explored for bio-active asymmetric Biginelli scaffolds.

Induced variability and assessment of mutagenic effectiveness and efficiency in sorghum genotypes [Sorghum bicolor (L.) Moench

Purpose Sorghum is an important cereal crop, cultivated for food, fodder and biofuel. Mutation breeding techniques are used to create genetic variability for qualitative and quantitative traits in crop plants. The purpose of this study was to create induced variability and estimate mutagenic effectiveness and efficiency of physical and chemical mutagens in two sorghum cultivars. Materials and methods Gamma rays (100, 200, 300 and 400 Gy, Co60 source, Bhabha Atomic Research Center, Mumbai, India), ethyl methane sulfonate (0.1%, 0.2%, 0.3%, and 0.4% EMS, Sigma-Aldrich, Bangalore, India) and their combinations were used to mutagenize 296B (rainy season) and Parbhani moti (post-rainy) cultivars. Morphological and yield traits were analyzed for enhanced variability in qualitative and quantitative traits across M2 and M3 generations. Results Based on the mutagenic sensitivity, lethal dose at 50% survivability (LD50) for both the genotypes was found to be 269–281 Gy in case of gamma rays and 0.32–0.33% for EMS. Based on reduced germination and survivability, mutagenic sensitivity was dose dependent and genotype independent. High frequency of chlorophyll mutations (albino, xantha, viridis, variegated and chlorina) was linearly correlated with dose in both the genotypes. Among the favorable mutants, dwarf and brown midrib were isolated from Parbhani moti population, which could be used in the cross breeding programs. A combined treatment, 100 Gy + 0.1% EMS showed high mutagenic effectiveness and efficiency. Enhanced genetic variation for quantitative traits as measured by wide range values and coefficient of variation was attributed to the effect of physical and chemical mutagens. Early flowering and high grain yield (24–49% increase over control) mutants were identified in M2 and validated in M3 generation in both genotypes. Conclusions This study has revealed wide genetic variability and better effectiveness and efficiency of the physical (300 Gy) and chemical mutagens (0.2%) and their combination (200 Gy + 0.2%) across two sorghum genotypes. Significant correlations identified between quantitative traits will help in better selection in the segregating generations.

Screening of an Ethyl Methane Sulfonate Mutagenized Population of a Wheat Cultivar Susceptible to Fusarium Head Blight Identifies Resistant Variants.

Fusarium head blight (FHB) primarily caused by Fusarium graminearum is a key disease of small grains. Diseased spikes show symptoms of premature bleaching shortly after infection and have aborted or shriveled seeds, resulting in reduced yields. The fungus also deteriorates quality and safety of the grain because of production of mycotoxins, especially deoxynivalenol (DON), which can result in grain being docked or rejected at the point of sale. Genetic host resistance to FHB is quantitative, and no complete genetic resistance against this devastating disease is available. Alternative approaches to develop new sources of FHB resistance are needed. In this study, we performed extensive forward genetic screening of the M4 generation of an ethyl methane sulfonate-induced mutagenized population of cultivar Jagger to isolate variants with FHB resistance. In field testing, 74 mutant lines were found to have resistance against FHB spread, and 30 of these lines also had low DON content. Subsequent testing over 2 years in controlled greenhouse conditions revealed 10 M6 lines showing significantly lower FHB spread. Seven and 6 of those 10 lines also had reduced DON content and fewer Fusarium-damaged kernels, respectively. Future endeavors will include identification of the mutations that led to resistance in these variants.

Effect of life stage and target tissue on dose-response assessment of ethyl methane sulfonate-induced genotoxicity.

In order to investigate the possibility that treatment age affects the genotoxic response to ethyl methane sulfonate (EMS) exposure, we dosed gpt-delta neonatal mice on postnatal days 1-28 with 5-100 mg/kg/day of EMS and measured micronucleus (MN) induction in peripheral blood and gpt gene mutation in liver, lung, bone marrow, small intestine, spleen, and kidney. The data were compared to measurements from similarly exposed adult gpt-delta mice. Our results indicate that the peripheral blood MN frequencies in mice treated as neonates are not substantially different from those measured in mice treated as adults. There were, however, differences in tissue-specific gpt mutation responses in mice treated with EMS as neonates and adults. Greater mutant frequencies were seen in DNA isolated from kidney of mice treated as neonates, whereas the mutant frequencies in bone marrow, liver, and spleen were greater in the animals treated as adults. Benchmark dose potency ranking indicated that the differences for kidney were significant. Our data indicate that there are differences in EMS-induced genotoxicity between mice treated as adults and neonates; the differences, however, are relatively small.

Characterization and fine mapping of a lesion mimic mutant (Lm5) with enhanced stripe rust and powdery mildew resistance in bread wheat (Triticum aestivum L.).

A novel light intensity-dependent lesion mimic mutant with enhanced disease resistance was physiologically, biochemically, and genetically characterized, and the causative gene was fine mapped to a 1.28 Mbp interval containing 17 high-confidence genes. Lesion mimic mutants are ideal for studying disease resistance and programmed cell death photosynthesis in plants to improve crop yield. In this study, a novel light intensity-dependent lesion mimic mutant (MC21) was obtained from the wheat variety Chuannong16 (CN16) by ethyl methane sulfonate treatment. The mutant initially developed tiny lesion spots on the basal part of the leaves, which then gradually proceeded down to leaf sheaths, stems, shells, and awns at the flowering stage. The major agronomic traits were significantly altered in the mutant compared to that in the wild-type CN16. Furthermore, the mutant exhibited a lesion phenotype with degenerated chloroplast structure, decreased chlorophyll content, increased level of reactive oxygen species, and increased resistance to stripe rust and powdery mildew. Genetic analysis indicated that the lesion phenotype was controlled by a novel single semi-dominant nuclear gene. The target gene was mapped on chromosome arm 2AL located between Kompetitive Allele Specific PCR (KASP) markers, KASP-4211 and KASP-5353, and tentatively termed as lesion mimic 5 (Lm5). The fine mapping suggested that Lm5 was located in a 1.28 Mbp interval between markers KASP-5825 and KASP-9366; 17 high-confidence candidate genes were included in this genomic region. This study provides an important foundational step for further cloning of Lm5 using a map-based approach.

Multivariate analysis of mutant wheat (Triticum aestivum L.) lines under drought stress

Drought is a major threat to wheat production worldwide and development of drought-tolerant wheat genotypes may improve the production of this cereal crop in drought affected regions. In the present study, 24 mutant lines were developed by treating a wheat variety 'NN-Gandum-1(NN-1) with ethyl methane sulfonate (EMS). The M5, M6, and M7 generations were evaluated for quantitative traits under well irrigated and rainfed conditions. Furthermore, best performing mutants of M6 and M7 generations were examined for biochemical parameters. A substantial correlation was found among the quantitative traits (plant height, spike length, tillers per plant, number of spikelets per spike and thousand grain weight) which confirmed their high heritability and relative relation. Principal component analysis (PCA) showed that the first two PCs with eigenvalues >1 had more than 50% of the total variability among the mutant lines under normal and water limited conditions. Cluster analysis depicted that considerable variations existed among all mutant lines. Among these mutant lines, some mutants showed significant variation consistently larger than corresponding values of wild type in multigeneration replication. Using drought tolerance indices (Stress tolerance index and geometric mean productivity), high yielding mutant lines under drought stress were selected. Across all these three-mutant generations, mutant lines NN1-M-363, NN1-M-506, NN1-M-700, NN1-M-701, and NN1-M-1621 showed significant improvement as compared to wild type in response to stress conditions. The selected five drought tolerant mutants showed increased accumulation of proline contents, total soluble sugars, total free amino acids, while decreased total chlorophyll content, carotenoids, and total soluble proteins. Results suggested that the morphological traits and yield attributes along with biochemical parameters could be utilized for the evaluation of wheat cultivars under drought stress.

Early flowering, good grain quality mutants through gamma rays and EMS for enhancing per day productivity in rice (Oryza sativa L.)

Purpose Rice is the prominent crop of world and it frames the important component of food chain. But, long duration of highly preferred rice varieties requires more water and are prone to biotic and abiotic stresses. Short duration rice varieties serve as a promising alternative in such cases. Hence, the present study was taken up to induce early flowering mutants in popular rice variety Bapatla 2231 (BPT 2231), a long duration medium slender rice variety using gamma rays and Ethyl Methane Sulfonate (EMS) treatment combinations. Materials and methods Five hundred well filled and viable grains of optimum moisture content (12%) were subjected to irradiation with 8 doses comprising 100 Gy, 200 Gy, 300 Gy, 350 Gy, 400 Gy, 450 Gy, 500 Gy and 600 Gy treatments. For combination treatments, the irradiated seeds of gamma rays were further treated with 30 mM concentrations of Ethyl Methane sulfonate (EMS). The irradiated population were raised up to M3 generation for the identification of early maturing mutants. The isolated early maturing mutants in M3 generation were characterized for phenotypic, biochemical and grain qualities. Results The frequency of desirable early maturing mutants was observed more in combination treatments than the individual treatments of gamma rays. A total of 34 early maturing mutants (M) with variation for grain quality and biochemical traits were obtained. The mutants M-1, M-3, M-5, M-32, and M-34 were high yielding with fine grain type. The mutants M-4, M-17, M-18, M-19, M-20, and M-26 were high yielding with bold grain type. The mutants M-22, M-23, M-24, M-27, and M-28 were high in nutrient content. The early mutants matured 40−50 days earlier than control. Conclusion In this variety, the combination treatments showed higher mutation frequency, mutagenic effectiveness and efficiency than the gamma rays in induction of both chlorophyll and viable mutants. This study revealed that the combination treatments were more effective in inducing early mutants than the gamma ray treatments alone. The early maturing mutants can be released as variety after evaluation under different yield trials and the mutants with increased nutrient content and varied grain quality can be utilized in hybridization/heterosis breeding programs for rice crop improvement.

In vitro propagation of six selected sugarcane mutant clones through leaf explants

Six mutant clones of sugarcane with high productivity have been produced through tissue culture techniques combined with mutations using gamma-ray irradiation and Ethyl Methane Sulfonate. The six mutant clones have been tested for stability in the field. They are proven to have high productivity and yields, so that they are very potential to be developed as superior varieties. To support the planting material sufficiency of these clones, an efficient propagation method was needed. Media formulations with different physical properties and composition of growth regulators were tested to obtain high seedling propagation rates. The media formulation for callus induction was Murashige dan Skoog (MS) + 3 mg/l 2,4-D + 3 g/l casein hydrolysate + 3% sucrose and for shoot regeneration was MS + 0,5 mg/l BA + 0,1 mg/l IBA + 100 mg/l PVP and 2% sucrose. Shoot proliferation was carried out on MS liquid (1, ½) + (0.3; 0.5 mg/l) BA + 0.1 mg/l IBA + 1 mg/l Kinetin + (0; 0.5 mg/l) GA3+ sucrose 2%. The results showed that callus induction, callus regeneration, and shoot proliferation of sugarcane mutant clones were influenced by the genotype and medium composition. The fastest callus induction was obtained from the MSP-4 clone (5.82 days), and the longest was MSB-7 (8.82 days). The largest callus diameter was obtained from MSB-6 clone on MS medium containing 1 mg/l BA, 100 mg/l PVP, and 2% sucrose. The highest number of shoots was obtained from the MSB-6 clone, while the least number of shoots conducted from the MSB-8 clone. The MSB-8 clones were more difficult to regenerate compared to the others. The best media formulation for shoot proliferation was ½ MS containing 0.5 mg/l BA, 1 mg/l Kinetin, and 0.1 mg/l IBA, while the best formulation for rooting was ½ MS.

Genotoxic effect of two commonly used textile dyes Reactive Blue 19 and Reactive Black 5 using Allium cepa L. as an indicator

In this study, the cytotoxic and genotoxic effects of Reactive Blue 19 and Reactive Black 5 were investigated using the Allium test and comet assay. These chemicals are fabric dyes used in textile industries in various parts of Turkey. Bulbs with roots of Allium cepa L. were treated with different concentrations (25, 50 and 100 ppm) of these textile dyes for 24 h. and 48 h. The root tips were processed for cytological studies by the aseto-orcein squash procedure. Distilled water and methyl methane sulfonate (MMS, 10 ppm) were used as an negative and positive control, respectively. Exposure of Reactive Blue 19 and Reactive Black 5 significantly decreased mitotic index values. Additionally, all treatments changed the frequency of mitotic phases when compared with the control groups. These dyes increased chromosome aberrations in test material. Among these abnormalities were anaphase bridges, c-mitosis, laggards, micronuclei and stickiness. A significant increase in DNA damage was also observed at all concentrations of both Reactive Blue 19 and Reactive Black 5 examined by comet assay.

Genotyping-by-Sequencing Based Genetic Mapping Identified Major and Consistent Genomic Regions for Productivity and Quality Traits in Peanut.

With an objective of identifying the genomic regions for productivity and quality traits in peanut, a recombinant inbred line (RIL) population developed from an elite variety, TMV 2 and its ethyl methane sulfonate (EMS)-derived mutant was phenotyped over six seasons and genotyped with genotyping-by-sequencing (GBS), Arachis hypogaea transposable element (AhTE) and simple sequence repeats (SSR) markers. The genetic map with 700 markers spanning 2,438.1 cM was employed for quantitative trait loci (QTL) analysis which identified a total of 47 main-effect QTLs for the productivity and oil quality traits with the phenotypic variance explained (PVE) of 10–52% over the seasons. A common QTL region (46.7–50.1 cM) on Ah02 was identified for the multiple traits, such as a number of pods per plant (NPPP), pod weight per plant (PWPP), shelling percentage (SP), and test weight (TW). Similarly, a QTL (7.1–18.0 cM) on Ah16 was identified for both SP and protein content (PC). Epistatic QTL (epiQTL) analysis revealed intra- and inter-chromosomal interactions for the main-effect QTLs and other genomic regions governing these productivity traits. The markers identified by a single marker analysis (SMA) mapped to the QTL regions for most of the traits. Among the five potential candidate genes identified for PC, SP and oil quality, two genes (Arahy.7A57YA and Arahy.CH9B83) were affected by AhMITE1 transposition, and three genes (Arahy.J5SZ1I, Arahy.MZJT69, and Arahy.X7PJ8H) involved functional single nucleotide polymorphisms (SNPs). With major and consistent effects, the genomic regions, candidate genes, and the associated markers identified in this study would provide an opportunity for gene cloning and genomics-assisted breeding for increasing the productivity and enhancing the quality of peanut.

Elevated particulate methanesulfonate, oxalate and iron over Sydney Harbour in the austral summer of 2019-20 during unprecedented bushfire activity

In the austral summer period, Dec 7, 2019 to Mar 1, 2020, which included unprecedented bushfire activity, total suspended particulate concentrations of methanesulfonate (MSA), oxalate, labile iron, SO42−, Cl−, Na+, K+, NH4+, water soluble solids (WSS) and water soluble organic carbon (WSOC) were measured from weekly filter samples at Seaforth on the northern edge of Sydney Harbour. Particle number concentrations (PNC) and meteorological conditions were also measured at hourly intervals. These entities were previously measured in the spring-summer period, Sep 30, 2018 to Feb 24, 2019. Data analysis is focussed on inter-comparison between the summer of 2018–19 (S1) and 2019–20 (S2) spanning the period 7 Dec to 24 Feb. During S2 seasonally averaged MSA, oxalate, labile iron, K+, PNC (0.5–2.5 μm), and WSOC were elevated by 102%, 57%, 64%, 72%, 108%, and 1907%, respectively, by comparison to S1. Aqueous phase secondary formation of oxalate and a marine source of MSA were evident during S1; however, during S2 it was indicated that the bushfires created conditions where oxalate formation was shifted away from aqueous phase processing and MSA became associated with bushfire generated WSOC aerosol. Correlation changes at the a = 0.05 significance level between S1 and S2 for MSA, and for oxalate, are examined and discussed in relation to effects apparently caused by the bushfires. It is indicated that the biogeochemical cycling of sulfur and carbon, and particulate deposition of iron micronutrient to the South Pacific Ocean, may be altered by bushfires in Australia.

İleri Generasyon Mercimek Mutant Genotiplerinin İmazamoks Herbisitine Toleranslarının Belirlenmesi

In Turkey, one of the essential grain legumes is lentil. It is usually perceived as a weak competitor with weeds. The research objective was to determine the tolerance of selected 145 mutagenized lentil genotypes at M5 generation to imazamox herbicide including 139 M5 lentil genotypes derived from Ethyl Methane Sulfonate (EMS) mutagenized seeds of cultivar Firat-87 and 6 control lentil cultivars were screened for imazamox herbicide tolerance. Experiments were carried out in the greenhouse and field. Herbicide was applied at 150% of the recommended dose of (100 ml/ha, or 40 g a.i/ha) imazamox when the plants were between 5 - 6 node stage. The response of the genotypes to the herbicide was evaluated by measuring the plant height as a sign of the growth and also by visual scoring of foliar damage with a 1 to 5 scale at 45 and 60 days after a spraying in the field experiment and at 30 and 60 days after a spraying in the greenhouse experiment. The genotypes were categorized based on their reactions to herbicides as highly tolerant, tolerant, moderately tolerant, sensitive, and highly sensitive. The results showed significant differences among the genotypes for tolerance to the herbicide. At 60 days after spray, most of the genotypes showed some of the recoveries in both experiments. Five genotypes (IMI-124, IMI-128, IMI-130, IMI-138, and IMI-139), displayed high herbicide tolerance in both experiments. The tolerant genotypes can be exploited in future breeding programs for improving herbicide tolerant lentil varieties.

Improvement and screening of astaxanthin producing mutants of newly isolated Coelastrum sp. using ethyl methane sulfonate induced mutagenesis technique

Natural astaxanthin is known to be produced by green microalgae, a potent producer of the most powerful antioxidant. To increase the productivity of astaxanthin in microalgae, random mutagenesis has been extensively used to improve the yield of valuable substances. In the presented work, a newly isolated Coelastrum sp. was randomly mutagenized by exposure to ethyl methane sulfonate and further screened using two approaches; an approach for high growth mutant and an approach for high astaxanthin producing mutant with a high-throughput screening method using glufosinate. Among these, mutant G1-C1 that was selected using glufosinate showed the highest of total carotenoids (45.48±1.5 mg/L) and astaxanthin (28.32±2.5 mg/L) production, which was almost 2-fold higher than that of wild type. This study indicates that random mutagenesis via chemical mutation strategy and screening using glufosinate successfully expedited astaxanthin production in a mutated strain of a Coelastrum sp.

Mutation breeding for heat and drought tolerance in tepary bean (Phaseolus acutifolius A. Gray)

Tepary bean (Phaseolus acutifolius A. Gray) is more heat and drought tolerant than common bean (P. vulgaris L.). Four hundred mutant lines of two tepary accessions (G40068 and G40159) were generated by ethyl methane sulfonate (EMS) treatment. In preliminary studies of the M5 mutant lines under abiotic stress, three mutant lines (CMT 38, CMT 109, CMT 187) were selected from six mutated lines based on morpho-physiological traits and superior yield and advanced to the M6 generation. The M6 mutant lines were uniform and genetically stable. These mutant lines and their original (M0) parents were evaluated for heat and drought tolerance under greenhouse conditions. Their performance was evaluated for morpho-physiological attributes, seed yield and yield components. Under high temperature and drought conditions, the CMT 38 mutant (M6 line) and its original tepary (M0) accession (G40068) showed greater values of pod biomass, pod number and 100-seed biomass than the other lines tested. The CMT 109 and CMT 187 mutant lines and their G40159 original accession (M0) also showed the highest value of seed number under high temperature and drought conditions. This suggests that the previous screening performed during the population advancement of these mutant lines, based on morphological traits like growth habit, was not detrimental to the yield variables evaluated here. Under combined heat and drought conditions, different parameters could be incorporated into tepary breeding programmes, as selection criteria to screen genotypes for tolerance to heat and drought stress. These parameters included: chlorophyll (SPAD) readings, seed biomass, 100-seed biomass and seed number because they explain the observed variance in the principal component analysis. Two additional traits (root biomass and stem diameter) were also identified as useful attributes, based on univariate analysis. The mutant lines evaluated here offer potential for further improvement of tepary bean to high temperature and drought

Characterization and Proteomic Analysis of Novel Rice Lesion Mimic Mutant with Enhanced Disease Resistance

Lesion mimic mutants (LMMs) are plants that spontaneously form lesions without pathogen infection or external stimulus and exhibit resistance to pathogens. Here, a rice LMM was created by ethyl methane sulfonate mutagenesis, named as hpil (hydrogen peroxide induced lesion). Diaminobenzidine and trypan blue staining showed that large amounts of H2O2 were produced and cell death was occurred at and around the parts of lesion mimic in the rice leaves. The phenotype of hpil is controlled by a single recessive gene, localized at a 2 Mb interval on chromosome 2. The data suggested that hpil is a novel LMM with enhanced bacterial and fungal disease resistance, and multiple pathogenesis-related proteins (PRs) were up-regulated. The proteomes of leaves at three positions (different degrees of lesion mimic severity) were characterized in hpil compared with its wild type plant. Differentially expressed proteins were detected by two dimensional difference gel electrophoresis and 274 proteins were identified by MALDI TOF/TOF™. These proteins were related to metabolic process, cellular process and response to stimulus, with mostly down-regulated in hpil leaves. Many of these proteins were related to the Calvin cycle, photosynthetic electron transport chain, glycolysis/gluconeogenesis and phosphonates pathways. Some resistance-related proteins including 14-3-3 proteins, OsPR10 and antioxidases such as peroxidase, superoxide dismutase and ascorbate peroxidase were up-regulated in leaves with lesion mimic. These results provide the foundation for cloning of the target gene and shed light on the mechanism involved in autoimmunity of rice.

Time dependent selected reaction monitoring based enhanced sensitivity approach for the determination of alkyl sulfonates and anisyl cyanides in Venlafaxine hydrochloride by Gas chromatography with mass spectrometry

Venlafaxine hydrochloride (VEN) is a newer generation antidepressant drug. Based on the synthetic procedure of VEN, formation of anisyl cyanides and alky sulfonates are possible. These impurities are known for their acute toxicity, come in the hazard and genotoxic class. Henceforth, it is necessity to control them in VEN at their acceptance level. Herein, sensitive t-SRM based gas chromatography coupled with triple-quadrupole mass spectrometry (GC–MS/MS) method has been developed for the trace level estimation of six potential impurities namely methyl methane sulfonate (MMS), ethyl methane sulfonate (EMS), isopropyl methane sulfonate (IPMS), o-anisyl cyanide (o-AC), m-anisyl cyanide (m-AC) and p-anisyl cyanide (p-AC); (alkyl sulfonates and anisyl cyanides) in VEN. The auto SRM tool was employed for the optimization of method parameters like SRM pair and collision energy to achieve the highest sensitivity. Various solvents as diluent were evaluated to optimize the extraction based recovery of all analyte. The method has been validated as per the ICH guideline (Q2R1). Obtained validation results proved the selectivity, sensitivity and robustness of the method. The correlation coefficient (r) values of all six impurities are more than 0.9988 in the range of 0.0003–0.6360 µg mL−1. Observed mean recoveries of all analytes are within the range of 93.21 to 97.77%. Method found to be capable to quantify of alkyl sulfonates and anisyl cyanides in the range of 4.5–24.1 ng g−1. Hence, the developed method is simple, direct, convenient, rapid and sensitive enough for its intended use in VEN trace impurity analysis.

Delaying or delivering: identification of novel NAM-1 alleles that delay senescence to extend wheat grain fill duration.

Senescence is a complex trait under genetic and environmental control, in which resources are remobilized from vegetative tissue into grain. Delayed senescence, or ‘staygreen’ traits, can confer stress tolerance, with extended photosynthetic activity hypothetically sustaining grain filling. The genetics of senescence regulation are largely unknown, with senescence variation often correlated with phenological traits. Here, we confirm staygreen phenotypes of two Triticum aestivum cv. Paragon ethyl methane sulfonate mutants previously identified during a forward genetic screen and selected for their agronomic performance, similar phenology, and differential senescence phenotypes. Grain filling experiments confirmed a positive relationship between onset of senescence and grain fill duration, reporting an associated ~14% increase in final dry grain weight for one mutant (P<0.05). Recombinant inbred line (RIL) populations segregating for the timing of senescence were developed for trait mapping purposes and phenotyped over multiple years under field conditions. Quantification and comparison of senescence metrics aided RIL selection, facilitating exome capture-enabled bulk segregant analysis (BSA). Using BSA we mapped our two staygreen traits to two independent, dominant, loci of 4.8 and 16.7 Mb in size encompassing 56 and 142 genes, respectively. Combining association analysis with variant effect prediction, we identified single nucleotide polymorphisms encoding self-validating mutations located in NAM-1 homoeologues, which we propose as gene candidates.

Biochemical and behavioral responses in carp fish exposed to tricaine methane sulfonate (MS-222) as anesthetic drug under transport conditions

This study aims to determine the effects of Tricaine methane‐sulfonate (MS-222) at concentration 150mg/l for one hour as an anesthetic agent to reduce the stress conditions during transfer the fish. Forty fish (Cyprinus carpio) were divide in to four groups: the 1st is control group (fish without both transfer and anesthesia), the 2nd group (fish transfer without anesthesia), the 3rd group (fish anesthesia without transfer) and the 4th group (fish transfer with anesthesia). The induction time reached to 3 minutes while the recovery time take at lasted for 7 to 10 minutes. Furthermore, the cortisol was significantly decreased in fish serum in both anesthetized groups that were transferred or that were left in the pond without transmission in compare to control groups (both transfer and without transfer fish). The glucose level rose significantly (P <0.05) in the serum of fish in both 3rd and 4th groups in compare to control groups. There was no difference in hematological parameters (hemoglobin and packed cell volume), blood sodium ion concentration, and enzyme activity represented by Alanine amino transferase in all groups while level of creatinine phosphate was decrease significantly in both groups 2nd and 3ed in compare to 1st group. It can be concluded that tricaine methane‐sulfonate stimulates the recovery and shortens the time of induction and reduces the stress condition caused by fish transport.

TPR domain coding gene ST2 may be involved in regulating tillering and fertility in rice

A decrease in the tiller number and male sterility will lead to a decline in the rice yield. Therefore, it is significant to study the molecular mechanism of controlling the tiller number and regulating the male reproductive development. The mutant st2 (single tiller 2) was induced by ethyl methane sulfonate (EMS) in the indica maintainer line Xinong 1B and showed single tillering and male sterility. I&lt;sub&gt;2&lt;/sub&gt;-KI staining showed that the st2 pollen was aborted. The scanning electron microscope (SEM) observation underlined that the anther of st2 became smaller, the wax of the epidermis reduced, the inner wall shrank and the Ubisch body decreased, the pollen collapsed, and the germination pore developed abnormally. The genetic analysis discovered that the trait was controlled by a single recessive nuclear gene located on chromosome 3. LOC_Os03g05540 encoding a tetratricopeptide repeat (TPR) domain was identified as the candidate gene by sequencing. The quantitative real-time polymerase chain reaction (qRT-PCR) analysis indicated that ST2 was highly expressed in the stem apical meristem (SAM) and the initial stage of meiosis during the anther development. The subcellular localisation indicated that ST2 is a nuclear and plasmic localisation protein. The homology analysis demonstrated that ST2 was evolutionarily conserved. These results laid a foundation for further study of the ST2 function.

Determining ethyl methane sulfonate-mediated (EMS) mutagenesis protocol for inducing high biomass yield in fodder barley (Hordeum vulgare L.)

Barley (Hordeum vulgare L.) has narrow genetic base for selection of promising ideotypes for the following traits: high biomass yield for livestock feed, enhanced agronomic and nutritional traits, and resistance to biotic and abiotic stresses. Ethyl methane sulfonate (EMS) mutagenesis offers opportunities for inducing genetic variation for key traits for development of feed barley ideotypes. The objective of this study was to determine optimal EMS dosage and exposure time to induce genetic variation for selection of high biomass yield six-row feed barley mutants. Five EMS dosages (i.e. 0.1%, 0.3%, 0.5%, 0.7% and 0.9% v/v) and five exposure times (i.e. 0.5 hr, 1 hr, 1.5 hr, 2 hrs and 2.5 hrs) were used on a six-row fodder barley using a 5 x 5 factorial treatment structure in a complete randomized design with three replications. Non-treated seed were used as a comparative control. Data was recorded for percent germination, seedling survival, shoot height, root height, shoot and root biomass. Significant (p &lt; 0.05) EMS dosage x exposure time was observed for assessed traits indicating their influence on phenotypic variation in feed barley. Overall, a declining trend was observed for assessed traits with increased EMS dosage and exposure time. The LD50 value of 0.64% (v/v) EMS dosage was identified as an optimal dose for large-scale mutagenesis protocol to select fodder barley mutants with high biomass yield